Multiomic approaches for assessing the role of natural microbial communities in nitrous oxide emission from Midwestern agricultural soils

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Anthropogenic activities such as fossil fuel consumption and industrial nitrogen (N) fixation processes have increased the N inputs into the environment. Even though the central role of microbes in the cycling of N is recognized, the identification and diversity of these microbial pathways in agricultural soils are still lacking. This scarcity of information limits the development of more accurate, predictive models of N-flux including the role of microbes in the generation and consumption of important nitrogenous greenhouse gases (e.g., nitrous oxide, N2O). The advent of new high-throughput nucleic acid sequencing technologies allows nowadays the exploration of soil microbial communities that were previously insufficiently studied based on cultivation and PCR approaches. In this work, we integrated experimental data and bioinformatics approaches to identify and quantify indigenous soil microorganisms participating in the cycling of nitrogen in agricultural fields, particularly those involved in the generation and consumption of N2O. We developed a new bioinformatic approach, called ROCker, to accurately detect target genes and transcripts in complex short-read metagenomes and metatranscriptomes, which offered up to 60-fold lower false discovery rate compared to the common strategy of using e-value thresholds. Using ROCker, we found an unexpectedly high abundance of nitrous oxide reductase genes, the only known biological sink of N2O, in soil and aquatic environments. In two particular soil types that typify the Midwest cornbelt, we show that microbial communities are remarkably stable across the year compared to other environments except during nitrogen fertilization events, which stimulate the activity of novel nitrogen-utilizing Nitrospirae and Thaumarchaeota taxa. Lastly, we assessed the predictive power of omic techniques in estimating nitrification process rates in incubated soil mesocosms and found high correlations between target transcripts and experimentally measured nitrification activity, providing new molecular means to measure microbial activity in-situ. These findings have implications for understanding the diversity and dynamics of natural microbial communities controlling the N cycle in soils.